The ability to distinguish self from non-self is essential in maintaining proper immune homeostasis. Most self-reactive T cells are eliminated in the thymus through negative selection;however some cells bearing T cell receptors that recognize self proteins may still escape thymic selection and enter the peripheral system as mature T cells. To avoid autoimmunity, those cells must be inactivated. One of the most important mechanisms involved in the maintenance of peripheral tolerance is suppression by regulatory T cells (Tregs). Natural Tregs are generated in the thymus and constitute 5-10% of the total CD4+ T cell population. Tregs control autoreactive T cells by suppressing cytokine expression and effector functions of T helper and cytotoxic T cells. Although the function of Tregs has been the object of intense research during the last years, the mechanisms responsible for the inactivation of T cells by Tregs remain yet poorly characterized. The overall goal of this proposal is to elucidate the molecular mechanisms underlying Treg mediated inactivation of effector T cell functions. Recent reports have indicated that members of the Nuclear Factor of Activated T cells (NFAT) may play a role in controlling self-reactive T cells by regulating the induction of anergy and the generation of Tregs. In this proposal we will determine if the interaction of Tregs with T helper cells may also result in the induction of a calcium/NFAT-dependent program that leads to the suppression of T cell activation. The role of the activation of NFAT proteins in Treg function will be studied in vitro and in vivo and the consequences of the upregulation of the expression of specific genes analyzed. A detailed understanding of the mechanisms that underlie Treg-mediated suppression of effector T cell function should provide valuable information to develop new therapies designed to induce tolerance in autoimmune disease and to prevent graft rejection, or to block Treg function during the treatment of cancer or chronic infections. PUBLIC HEALTH RELEVANCE Regulatory T cells have been identified as key regulators in the control of immune reactions against our own tissues. Their function is also crucial to understand how chronic infections and cancer cells can evade effective immune responses. The mechanisms that underlie the function of these cells are yet poorly understood. The aim of this project is to characterize how regulatory T cells mediate inhibition of other active T cell populations. The information obtained from this project will be very valuable for the design of new therapeutic approaches to control regulatory T cell function in the treatment of diseases such as autoimmunity, graft rejection and cancer.